The present application is a national phase application of international application Ser. No. PCT/JP98/01972, filed under 37 C.F.R. xc2xa7371, which claims benefit of priority of JP 113126/1997. These applications are explicitly incorporated herein by reference in their entirety and for all purposes.
The present invention relates to a method for detecting or predicting ischemic disorders by using as an indication the concentration of human lipocalin-type prostaglandin D synthase (hereinafter, simply referred to as xe2x80x9chPGDSxe2x80x9d) in body fluid samples.
Prostaglandin D2 is a physiologically active substance synthesized from arachidonic acid, and can be produced from prostaglandin H with PGD synthase. Recently, it has been found that hPGDS is identical with xcex2-trace that has been known to be found in human cerebrospinal fluid (hereinafter, simply referred to as xe2x80x9cCSFxe2x80x9d) in a large amount and such distribution of hPGDS in a human body suggests some association with neurological disorders. hPGDS has also been known to be found in semen, amniotic fluid and the like, and therefore it has been proposed to utilize hPGDS for the assessment of fertility (i.e., assessment of azoospermia), diagnosis of fetal development, and the like. However, no report suggesting the correlation between hPGDS and ischemic disorders has been found.
Arteriosclerosis, which is a major factor of ischemic disorders, has two types: atherosclerosis arising at relatively large arteriae and aortae; and arteriolosclerosis arising at peripheral arteriolae. Atherosclerosis provokes outgrowth of foam cells, proliferation of fibrae, hyperplasia of tunica intima vasorum, caused by lipid deposits, thrombogenesis, and calcification resulting in stenosis or occlusion of lumens; thus ischemic disorders are produced. Arteriolosclerosis provokes the increase in connective fibrae in tunica intima, resulting in strong hyalinization. Such hyalinization often extends to tunica media. In this case, the arteria are hyalinized and thickened and lumens are narrowed, resulting in ischemic disorders. In general, the cause of arteriosclerosis is considered as follows. When endothelial cells are impaired by any factor such as hyperlipidemia or hypertension, monocytes infiltrate into the walls of the arteriae from the circulating blood together with serum components and were transformed to macrophages. The macrophages incorporate oxidized low density lipoprotein (LDL) that had been undergone oxidization and denaturation in the vessel walls and were transformed to foam cells. The activated endothelial cells, macrophages or adhesive platelets release growth factors or chemotaxis factors, which induces the migration into tunica intima or proliferation of smooth muscle cells. The smooth muscle cells also accumulate lipids therein and consequently are transformed to foam cells. Thereafter, hyperplasia of tunica intima, atheroma formation, stenosis of lumen, ulcer formation and cicatrization are developed. In this manner, arteriosclerosis plaques are formed.
Disorders in which interference with blood flow is induced by coronary sclerosis are called ischemic heart disorders, such as angina pectoris and myocardial infarction. Ischemic heart disorders provoke cerebral circulatory disorders (e.g., cerebral infarction) at cerebral arteriae and obturating arteriosclerotic disorders at peripheral arteriae. Arteriolosclerosis is accompanied by hyperplasia and degeneration of tunica intima and tunica media caused by hyalinization thereof, and it predominantly provoke cerebral and renal disorders.
All types of ischemic disorders progress without any of their characteristic symptoms or signs at the early stage, and none of the clinical symptoms appears until the lesion progresses to an advanced stage. However, up to now, no effective biochemical detection method applicable for the prediction of ischemic disorders during such period of stage has been developed. At the point of time where the symptoms appear and abnormality in the results of clinical trials associated with the development of the symptoms are observed, the lesions do give rise to the terminal phase. At present, there is no screening method that exactly diagnoses the lesions at the early stage, therefore diagnosis of arteriosclerosis is based on the cumulative assessment of risk factors such as lipids in serum, hypertension and smoking.
As mentioned above, there is no reliable method for diagnosis of the lesion at the early stage for ischemic disorders. Moreover, after any clinical symptom appears, patients with ischemic disorders are needed for extremely severe life management and therapy. Therefore, it is an urgent demand to develop a screening method that exactly diagnoses the lesion of an ischemic disorder at its early stage.
Prostaglandin D2 exhibits supressive action against organic change associated with arteriosclerosis, such as an inhibiting coagulation. Therefore, the present inventors have focused on the correlation between prostaglandin D2 synthesizing enzyme, hPGDS, and arteriosclerosis. For the purpose of overcoming the above-mentioned problems, the present inventors determined the hPGDS concentrations in body fluid samples (e.g., blood, cerebrospinal fluid, urine) from normal subjects, and established reference value for each type of body fluid sample. Based on the reference value, it was examined whether the hPGDS concentration in a body fluid sample from a test subject was suitable as the indication of the early stage of an ischemic disorder, in other words, whether the hPGDS concentration in a body fluid sample has any correlation with the ischemic disorder. As a result, it was found that the hPGDS concentration in fluid samples collected from a patient with ischemic disorder or body fluid sample from the patient collected before he was diagnosed as such showed significant difference from the reference values, hPGDS concentrations established from normal subjects. This finding leads the possibility of prediction or detection of an ischemic disorder in a patient at the early stage by monitoring the kinetics of hPGDS in a body fluid sample. It is also found that the method of the present invention can exactly predict an ischemic disorder at its earlier stage, prior to the prediction method based on the cumulative assessment of risk factors (e.g., hypertension, lipids in serum, smoking).
Accordingly, the present invention provides a method for detecting or predicting ischemic disorders comprising determining the concentration of hPGDS in a body fluid sample. The body fluid sample includes blood, urine, cerebrospinal fluid, saliva or semen. The method for the determination of the hPGDS concentration includes an immunological assay. In the present invention, it is preferable that the detection or prediction of an ischemic disorder be made by the comparison between the hPGDS concentration in a body fluid sample from a test subject and those from a normal subject.
The ischemic disorder to be detected or predicted includes those resulted from arteriosclerosis or embolus; ischemic heart disorders (e.g., myocardial infarction, angina pectoris); cerebral infarction; intracranial hemorrhages (e.g., intracerebral hemorrhage, subarachnoid hemorrhage, subdural hemorrhage); aneurysms (e.g., dissecting aneurysm, abdominal aortic aneurysm); nephrosclerosis; myocardial infarction developed as a sequela of Kawasaki disease; and the like.
The present invention also provides a kit for detecting an ischemic disorder which comprises an antibody specific to hPGDS. Preferably, the kit comprises first and second antibodies specific to hPGDS. In this case, the second antibody is preferably capable of binding to a conjugate of hPGDS with the first antibody. Both the first and second antibodies are preferably monoclonal antibodies. An example of the first antibody is a monoclonal antibody 7F5 (described below), and an example of the second antibody is a monoclonal antibody 1B7 (described below).